Analysis of mutant platelet-derived growth factor receptors expressed in PC12 cells identifies signals governing sodium channel induction during neuronal differentiation

The mechanisms governing neuronal differentiation, including the signals underlying the induction of voltage-dependent sodium (Na⁺) channel expression by neurotrophic factors, which occurs independent of Ras activity, are not well understood. Therefore, Na⁺ channel induction was analyzed in sublines of PC12 cells stably expressing platelet-derived growth factor (PDGF) β receptors with mutations that eliminate activation of specific signaling molecules. Mutations eliminating activation of phosphatidylinositol 3-kinase (PI3K), phospholipase C(γ) (PLC(γ)), the GTPase-activating protein (GAP), and Syp phosphatase failed to diminish the induction of type II Na⁺ channel α-subunit mRNA and functional Na⁺ channel expression by PDGF, as determined by RNase protection assays and whole-cell patch clamp recording. However, mutation of juxtamembrane tyrosines that bind members of the Src family of kinases upon receptor activation inhibited the induction of functional Na⁺ channels while leaving the induction of type II α-subunit mRNA intact. Mutation of juxtamembrane tyrosines in combination with mutations eliminating activation of PI3K, PLC(γ), GAP, and Syp abolished the induction of type II α-subunit mRNA, suggesting that at least partially redundant signaling mechanisms mediate this induction. The differential effects of the receptor mutations on Na⁺ channel expression did not reflect global changes in receptor signaling capabilities, as in all of the mutant receptors analyzed, the induction of c-fos and transin mRNAs still occurred. The results reveal an important role for the Src family in the induction of Na⁺ channel expression and highlight the multiplicity and combinatorial nature of the signaling mechanisms governing neuronal differentiation.